Proper regulation of uterine blood flow during pregnancy is important not only for the growth and survival of the fetus, but also for cardiovascular well-being of the mother. Chronic hypoxia during the course of pregnancy is one of the most common insults to the maternal cardiovascular system and fetal development, and is associated with an increased risk of preeclampsia and fetal intrauterine growth restriction. Previous studies have demonstrated that chronic hypoxia has profound effects on uterine artery reactivity, and inhibits pregnancy-induced changes in uterine artery contractility. Normal pregnancy is associated with decreased uterine vascular tone and increased contractile reactivity to nonsynaptic alpha1-adrenergic-stimulation, due probably in part to a transient and reversible sympathetic denervation of the uterine artery during pregnancy. We have discovered recently that pregnancy increases the effect of extracellular signal-regulated kinase (ERK)in suppressing protein kinase C (PKC)-mediated contractions of the uterine artery. PKC has been proposed to play an important role in regulating the sustained contraction of smooth muscle, and hence, vascular tone. In addition, PKC produces a negative feedback regulation on alpha1-adrenoceptor-mediated contractions in vascular smooth muscle. These findings lead to the proposed studies of mechanisms, testing the central hypothesis that pregnancy upregulates the ERK pathway and downregulates the PKC pathway in uterine artery smooth muscle resulting in decreased basal vascular tone and enhanced action of alpha1-adrenoceptors, and this adaptation is inhibited by chronic hypoxia. To test this hypothesis, we propose a series of experiments in the uterine arteries from nonpregnant and near-term (approximately 140 days) pregnant sheep maintained near sea level (approximately 300m) and at high altitude (3,820 m) from 30 days gestation. The specific aims are:1. To test the hypothesis that pregnancy upregulates the ERK pathway resulting in downregulation of the PKC pathway in the uterine artery, and this adaptation is inhibited by chronic hypoxia, we will determine: 1.1 effects of ERK inhibition on PKC isozymes expression and activities; 1.2 effects of PKC activation and inhibition on ERK tyrosine phosphorylation and activities; 1.3 effects of cortisol on ERK and PKC isozymes expression and activities. 2. To test the hypothesis that pregnancy alters the ERK/PKC pathways resulting in a decrease in baseline myofilament Ca 2+ sensitivity and basal vascular tone, and this adaptation is inhibited by chronic hypoxia, we will determine the effects of activation and inhibition of PKC and/or ERK on: 2.1 Ca2+-stimulated increases in myosin light chain phosphorylation (LC20-P) and force in permeabilized tissues; 2.2 agonist-stimulated increases in intracellular free Ca 2+ concentration ([Ca2+]i), LC20-P, and force in intact tissues; 2.3 protein concentrations and phosphorylation levels of caldesmon, CPI-17, and MYPT1. 3. To test the hypothesis that pregnancy alters the ERK/PKC pathways resulting in an enhanced pharmacomechanical coupling of alpha1-adrenoceptors, and this adaptation is inhibited by chronic hypoxia, we will determine the effects of activation and inhibition of PKC and/or ERK on: 3.1 alpha1-adrenoceptor density and agonist binding affinity; 3.2 coupling efficiency of alpha1-adrenoceptor/inositol 1,4,5-trisphosphate (IP3) synthesis and tissue contractile sensitivity to IP3; 3.3 IP3 receptor affinity and density; 3.4 alpha1-adrenoceptor-mediated increases in [Ca2+]i. The results of the proposed studies will provide important original insights into biochemical, cellular, and physiologic adaptation mechanisms involved in adjusting uteroplacental circulation in response to pregnancy, and will help improve our understanding of problems associated with the maladaptation and abnormal pregnancy caused by chronic hypoxia and permit us to address them in a more meaningful way.